Pain is a non trivial sensation used as a survival mechanism by which organisms are made aware of the presence of harmful stimuli in their environment. Transient Receptor Potential Vanilloid-1 (TRPV1), a polymodal non-selective cation channel, has emerged as a key player in pain perception. TRPV1 is activated by noxious heat (>420C), protons, and vanilloid ligands such as capsaicin, the spicy hot component in chili peppers. Similar to voltage-gated potassium (Kv) channels, TRPV1 forms tetramers with each protomer consisting of six predicted transmembrane segments (S1-S6) with large cytosolic N- and C-termini. The S1-S6 segments are subdivided into the S1-S4 domain, known to interact with vanilloids, and the S5-P-loop-S6 pore domain. TRPV1 activation by vanilloids has been the subject of extensive research in the last decade. Nevertheless, the molecular mechanism(s) by which these compounds activate TRPV1 is not well understood. Hence, this proposal seeks to elucidate the molecular details as to how vanilloids interact with the S1-S4 domain and how conformational changes in this region are coupled to channel gating. To address these questions, I propose a multi-pronged approach gathering information from both biochemical and structural biology methods. Knowledge gained about TRPV1 activation by vanilloid-ligands will be useful for the development of TRPV1 agonists and/or antagonists with potential uses as therapeutics for the treatment of pain associated with a variety of health diseases.